Matrix-assisted laser desorption/ionization mass spectrometry analysis and thiol-group determination of isoforms of bovine cytochrome c oxidase, a hydrophobic multisubunit membrane protein.
Matrix-assisted laser desorption/ionization-mass spectra (MALDI-MS) are obtained from entire bovine heart (H) and liver (L) cytochrome c oxidase membrane protein complexes. Molecular masses of most of the subunits are in excellent agreement with the published sequences. Some corrections are necessary for the nuclear coded subunit IX, which is N-acetylated, and X, with a corrected C-terminal peptide sequence. The mass values of two of the three tissue-specific subunits (VIII-L and X-L) are not in agreement with the DNA-deduced sequences and have been corrected by protein sequencing. For the investigation of the cysteine status 7-diethyl-amino-3-(4'-maleimidylphenyl)-4-methylcoumarin proved to be an excellent site-specific reagent. MALDI-MS with the SH-reacted enzyme indicates disulfide bridges only in subunit VII and a distorted tetrahedral S coordination of the zinc in subunit VI.
Marx
M K
MK
Institut für Biochemie, Klinikum der RWTH Aachen, Germany.
Mayer-Posner
F
F
Soulimane
T
T
Buse
G
G
D000818:Animals; D002417:Cattle; D003576:Electron Transport Complex IV / Q000032:analysis* / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D007425:Intracellular Membranes / Q000201:enzymology; D007527:Isoenzymes / Q000032:analysis* / Q000737:chemistry; D008565:Membrane Proteins / Q000032:analysis* / Q000737:chemistry; D008929:Mitochondria, Heart / Q000201:enzymology; D008930:Mitochondria, Liver / Q000201:enzymology; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000032:analysis* / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013438:Sulfhydryl Compounds / Q000032:analysis*
D007527:Isoenzymes; D008565:Membrane Proteins; D009097:Multienzyme Complexes; D013438:Sulfhydryl Compounds; D003576:Electron Transport Complex IV